Elimination of unwanted and harmful proteins is of key importance for the function of a cell and an organism. Work from many laboratories including ours in the past decade have uncovered a large number of structurally related protein complexes, named cullin- RING E3 ubiquitin ligases. These E3 complexes function as "molecular death trigger'by tagging target proteins with a chain of death-tag, known as ubiquitin, thereby directing their destruction. Defective regulation of these pathways is manifest in human diseases including cancers. Our long-range goal is to understand the precise functioning of cullin-RING E3 type of "molecular death triggers," and initiate efforts to develop therapeutic strategies against human cancers. The short-term objective of our project is to determine how one such type, the CUL7 E3, functions to eliminate a protein target called IRS-1, thereby contributing to tumor suppression. Specifically, we will carry out biochemical experiments to identify the "built-in" code in IRS-1 that directs its molecular interactions with the CUL7 E3, which triggers ubiquitin tagging and hence degradation. We will define those cellular components that modify the IRS-1 degradation "code," setting it "ON" or "OFF." In addition, we will perform in- depth molecular and cellular analysis to understand how the CUL7 E3 is assembled into a "molecular death trigger." We will devote our major efforts to explore our preliminary findings that elimination of IRS-1 by the CUL7 E3 helps a cell suppress cancer by inactivating a major growth- promoting activity known as PI3K. Cell transformation assays will be developed to address this issue. In addition, we have recently discovered that IRS-1 could prevent cancer by driving cancerous cells to a permanent growth arresting state (senescence). Given its significant ramification in understanding cancer initiation and prevention, we will dissect the components of this novel senescence program and understand how it functions using molecular and cell biology tools. PUBLIC HEALTH RELEVANCE: This project intends to explore two anti-cancer activities that are executed by the CUL7 E3 ubiquitin ligase complex, initially discovered in this laboratory. Experiments are outlined to understand how the CUL7 E3 directs the degradation of the IRS-1 signaling protein, and how this process suppresses cancer by inactivating the PI3K-growth promoting function. Moreover, in-depth molecular and cell biology methods will be employed to explore the novel function by IRS-1 in inducing permanent cell arrest (senescence), which has emerged as a prominent anti-cancer mechanism in multi- cellular organisms including humans.